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Development of analytical methods for determination of phosphorylated components of bacterial cell membranes
Mikulecká, Jana ; Čabala, Radomír (advisor) ; Feltl, Ladislav (referee)
Phospholipids are dominant components of bacterial cell membranes, where they create double layers. Bacteria differ in their phospholipid composition determination of which can help in identification of important groups of microorganisms. Phospholipid composition of bacteria is influenced by many environmental factors, therefore its variation can be observed within one bacterial stem also. Because of its simplicity, thin layer chromatography is usually applied to identification and determination of bacterial phospholipids. Disadvantage of this method are the high demands of time, carefulness and skills of the analytical personnel. The increasing interest in the phospholipid double-layer promotes the detailed investigation of their fatty acid composition because the more detailed analyses allows for more information yield about bacteria. Gas chromatography hyphenated with mass spectrometry seems to be the best choice for these purposes. Fatty acid identity and total fatty acid content in phospholipid molecules could be determined by this method. Additionally, number, position and isomerism of double bonds and presence of other functional groups on hydrocarbon chain could be determined. Whereas a suitable and...
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Electrochemical determination of chlortoluron using carbon paste electrode
Houšková, Lucie ; Zima, Jiří (advisor) ; Vyskočil, Vlastimil (referee)
Chlortoluron is a herbicide from the group of substituted ureas and it is used for weed control in production of cereals and poppy. Substituted ureas are widely used but these substances are known for their high mobility in soil, persistence and contamination of underground water. Therefore, the development of analytical methods suitable for residual concentrations determination is necessary. This thesis deals with the determination of chlortoluron using carbon paste electrode (CPE) by the methods of differential pulse voltammetry and high-performance liquid chromatography with electrochemical (HPLC-ED) and spectrophotometric (HPLC-UV) detection. The aim of the work was to find optimal conditions suitable for chlortoluron determination. Further, the possibility of using miniaturized carbon paste electrode (mCPE) and usability of tested methods for determination of chlortoluron in model samples of river water and soil were studied. Concentration dependences in deionized water were measured by the method of DPV in the Britton-Robinson (BR) buffer (pH = 3) and methanol (10 %, v/v). The limit of detection was 3,7∙10-7 mol∙dm-3 in the case of CPE and 8,7∙10-8 mol∙dm-3 in the case of mCPE. Also, concentration dependences were measured using HPLC with mobile phase containing BR buffer (pH = 4) and...
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Comparison of the fruits of some honeysuckle varieties in terms of content selected biologically active compounds
Veselý, Ondřej ; Diviš, Pavel (referee) ; Vespalcová, Milena (advisor)
This thesis is focused to the analysis of selected varieties of honeysuckle kamtschatica (Lonicera caerula var kamtschatica) and their comparison in terms of the content of biologically active substances. There were analysed only varieties grown in 2014. Theoretical part elaborates and characterized various parameters of biologically active compounds such as phenolic, anthocyanins and ascorbic acid and possibilities of their determination. Further it briefly surveys Kamchatka honeysuckle, its taxonomic and morphological properties and its nutritional properties. The experimental part of the work was focused on the description of the methods used for the analysis of biologically active substances and their analysis. Polyphenolic compounds and anthocyanin’s were determined spectrophotometrically and vitamin C was determined by high-performance liquid chromatography. The results and varieties were compared with each other. Overall, we can say that the variety with the highest content of all three analysed biologically active substances is a variety Valchová, which had the second highest content of polyphenolic compounds (546.3 mg · 100 g-1 fruits), the third highest contend of ascorbic acid (35.1 mg · 100 g-1 fruits) and the sixth highest content of anthocyanin (170.8 mg · 100 g-1 fruits)
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Analysis of drinks sweetened with stevia extracts
Procházka, Václav ; Hrstka, Miroslav (referee) ; Vespalcová, Milena (advisor)
The steviolglycosides are the natural, sweet substances from Stevia rebaudiana Bertoni. It affect human health positively and its sweetness is 300x stronger than the sweetness of sucrose. That's why it's used to sweetening the commercial products. Because of its potential properties it's good to have an appropriate method to isolate it. High performance liquid chromatography ( HPLC) is based on the separation of analyte between two immiscible phases with high pressure pump and appropriately chosen stationary phase in the column. Than the analytes come out from the column in different retention times. This master´s thesis follows up selection of the best HPLC system for isolation of the main steviolglycosides and its analysis in commercial products. In the theoretical part of the thesis is described the origin, basic characteristics, botanical description, cultivation and affect to the human health of Stevia rebaudiana Bertoni and its use in food industry. There are also concisely characterized the sweet substances contained in the plant, so called steviolglycosides. Than there are given the theoretical basics of high performance liquid chromatography, HPLC instrumentation and its specific applications at sleviolglycosides with the basic chromatographic parameters. The object of the first experimental part was to research the optimum conditions for time and separation effective chromatographic analysis and select the best chromatographic system for isolation of steviolglycosides. In the second experimental part, I have compared and defined the main steviolglycosides (stevioside, rebaudioside A) in nine selected products, commercially available in Czech republic, with the best chromatographic method. In these products was the contain of the stevioside or rebaudioside A confirmed or refused.
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Comparison of content anthocyanin pigments in selected cultivars of black elderberry and grape
Becková, Eliška ; Vítová, Eva (referee) ; Vespalcová, Milena (advisor)
The purpose of this thesis is to compare the contents anthocyans in different varieties of European elderberry (Sambucus nigra L.). It was also studied the relationship between the content anthocyans in selected samples of blue grape varieties (Vitis vinifera L.) and position of the track, where the grapes were grown. The theoretical part describes elderberries and grapes in terms of botanical, chemical, and in terms of their using in the food industry. The dominant part of this chapter is to describe anthocyans and possibilities for their determination in the elderberry and grapes. Emphasis has been placed on the pH-differential method and determination by high performance liquid chromatography. The experiments are divided into two parts. The aim of the first experimental part is comparison of the total anthocyanins content in fifteen varieties of elderberry. The performed analysis shows that in terms of anthocyanins content for cultivation of elderberry in the Czech Republic the most suitable varieties are Mammut, Samyl, Reis aus Vossloch and Haschberg. In the second part of the experiments were determined total anthocyanins content in nine varieties of blue grape grown in the Mikulov region. For determination were used HPLC and pH-differential methods, as well as in the first part. The highest content of pigments was found in varieties Alibernet and Neronet. The present study focused on the suitability of the location pointed to vineyard south to southeast orientation plantings. At the end of the experimental part is a comparison of both methods of determination. Method using high performance liquid chromatography to provide very accurate results compared to less demanding pH-differential method of determination. This is recommended only for tentative determination of anthocyanins content in real samples.
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Analysis of Echinacea content compounds
Michalíček, Jan ; Vespalcová, Milena (referee) ; Zemanová, Jana (advisor)
Rostliny rodu Echinacea jsou léčivé byliny stimulující imunitní systém. Používají se zvláště v boji proti chřipce a infekcím horních cest dýchacích. V této práci bylo analyzováno 7 komerčních výrobků obsahujících výtažky z rostliny Echinacea vysokoúčinnou kapalinovou chromatografií. Bylo provedeno kvantitativní stanovení derivátů kyseliny kávové, jež jsou látky zodpovědné za léčivé účinky rostliny. Zjištěné obsahy kyseliny chlorogenové, kávové, cichorové a echinakosidu byly porovnány v jednotlivých výrobcích. Dále byly připraveny extrakty z kořene, listů a květů třapatky nachové pomocí extrakce za zvýšeného tlaku extrakčního rozpouštědla při různých podmínkách, aby se z obsahu účinných látek v nich přítomných zjistilo, které extrakční podmínky jsou nejlepší. Z naměřených dat bylo zjištěno, že největší celková koncentrace obsahových látek je přítomna ve výrobku Imunit Echinaceové kapky firmy Simply You. Jako nejlepší extrakční podmínky se ukázaly teplota 80 °C v kombinaci s rozpouštědlem ve složení EtOH/voda = 40/60, a to jak při extrakci kořene, tak i listů a květů.
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Evaluation of glutathione content in plants as a marker of heavy metals environmental contamination
Borková, Marie ; Stoupalová, Michaela (referee) ; Opatřilová,, Radka (advisor)
Dependence of glutathione concentration on the amount of thallium in the plant was studied. Observed plant was maize (Zea mays) which was divided to two parts – root and overground. Two culture procedures were elaborated where seeds and young seedlings were cultivated in a solution of thallium of concentration 0, 1, 3, 5, 8, a 10 µmol/l. Extraction agents used during extraction were phosphate buffer and solution of ascorbic acid. Determination of glutathione was realized by capillary electrophoresis (CE) and high performance liquid chromatography (HPLC). Diode array detector (DAD) was used in both methods. Quantification of the thallium amount in the plant was done by method of inductively coupled plasma-atomic emission spectrometry (ICP-AES).
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The stability of biologically active compounds in elderberry (Sambucus nigra L.) inflorescence during processing
SUCHÁ, Renata
This thesis examines the stability of rutin in the inflorescences of the elder during processing. Elder (Sambucus nigra L.) is a medicinal plant that was used for medical purposes as early as in the Middle Ages. Both flowers (Flos Sambuci) and fruit (Fructus Sambuci) of the elder are on the list of official medicinal products of the State Institute for Drug Control. The main active phenolic substance is flavonoid rutin. Flavonoids are phenolic substances of plant origin. Among the best known properties of these substances is their antioxidant activity. They protect the low density blood lipoproteins from the oxidation modifications that cause the development of arteriosclerosis and hypertension. As a medical plant, the elder is used especially for the preparation of a medicinal drink. It is also commonly used as a material for the preparation of syrup. This product serves as a basis for a refreshing and tasty drink and lately is has also started to be sold in shops. For comparison, five single-flower syrups from the elderflower by different producers were bought. A method of preparation of the elderflower syrup was devised, based in one case on frozen elder inflorescences and in the other on dried inflorescences. The plant material came from wildly growing shrubs and it was collected in June 2013. Altogether, 8 syrups were prepared in the laboratory, 4 of them from fresh material and 4 from dried material. Two different methods were used. The first one was based on extraction using sugar solution, the second one on extraction using water solution where sugar was added afterwards. Another added substance was ethanol. The method for measuring the amount of rutin in the samples was a combination of two extraction steps. The first step was the usual exraction using aqueous methanol and the second step was extraction using a solid sorbent (SPE). The analysis of fresh and dried material concluded that dried elderflowers contain many times more rutin than fresh elder. Among the commercially produced syrups, the "Kitl Syrob Bezový" contained most rutin; a little less rutin was found in "Hamé Bylinný sirup Bezový květ". The concentration of rutin in the syrups prepared in the laboratory was much higher than in the commercial products. By using different methods of syrup preparation and their subsequent analysis, it was found that the syrups prepared from the dried inflorescences contained a higher amount of rutin than the syrups produced in the same way, but from frozen material. Addition of a small amount of ethanol had no effect on extraction of rutin from inflorescences. Free quercetin, which indicate a decomposition of rutin, wasn´t found in any of produced syrups. The produced syrups were used for the preparation of a drink and then underwent sensory analysis. The paired two-sided preference test included pairs of drinks consisting of a fresh elder drink and dried elder drink. The results showed that the evaluators preferred syrups from dried elderflowers in two of the cases, while in the other two evaluated pairs, the results were not statistically significant. In all cases, the evaluators most often found small and middle differences. The gained results show that the best method of syrup preparation uses dried plant material and sucrose solution as the extraction agent. The syrup prepared in this way contained most rutin and at the same time showed the smallest losses of this substance, which is beneficial to human health. Furthermore, it also tasted better in comparison with syrup prepared in the same way, but from fresh material.
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Content of selected phenolic compounds in spice plants.
BERANOVÁ, Zuzana
This work concerns the amount of certain phenols in some of the representatives of families Alliaceae, Lamiaceae and Apiaceae. Phenols in plants are widely represented and highly concentrated. Flavonoids are one of the smallest, yet quite significant classes of phenols. Ingestion of food containing flavonoids can prevent certain diseases such as Arteriosclerosis. cardiovascular and tumor diseases. This work focuses, for their special biological effects, on five flavonoids: Kaempferol, Quercetin, Myricetin, Apigenin and Luteolin. For determining the content of phenols a method of High-performance liquid chromatography (HPLC) was used. Three representative of the family Alliaceae, two representatives of the family Lamiaceae and three representatives of the family Apiaceae were analysed. Only edible parts of the plants were used for the analysis and the amount of phenols was compared in certain plants planted in beddings to the plants planted in greenhouses. At first, the qualitative representantion of phenols was ascertained by the HPLC method. The result is chromatographic profiles, which were then used in calculating the amount of particular phenols. Then the total amounts of kaempferol, quercetin, myricetin, apigenin and luteolin were found out thanks to HPLC method. The biggest amount of total kaempferol was determined in petroselium hortense planted in beddings (588 mg/kg of fresh sample) and in a greenhouse (340 mg/kg of fresh sample). The biggest amount of quercetin was determined in red onion planted in a greenhouse (773 mg/kg of fresh sample) and in ocimum basilicum planted in beddings (535 mg/kg of fresh sample). The biggest amount of apigenin was determined in petroselium hortense planted in a greenhouse (1790 mg/kg of fresh sample) and in petroselium chrispum planted in beddings (3690 mg/kg of fresh sample).
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The content of selected fenolic compounds in medicinal plants.
KREJČÍ, Zuzana
The work has been inquired into the problem of content determination of phenolic substances in medicinal plants traditionally used in Czech Republic. Phenolic substances belong to a group of natural compounds, which are purely plant origin. Flavonoids are a part of this extensive group of compounds. As for flavonoids, most attention is paid to quercetin and rutin. It is caused by their easy availability and very significant biological activity. These compounds embody a lot of positive biological effects. They have expressive antioxidant properties, inhibit lipid peroxidation, scavenge free oxygen radicals and bond into the chelates they inactivate some prooxidant metal ions. The latest researches have shown that thanks to their properties natural flavonoids can occurence of chronical diseases, such as arterosklerosis, cardiovascular or tumor disorder. Flavonoids are exploited both in traditional and modern medicine. Same other studies have evidenced that valuable sources of these biologically effective substances are traditionally used medicinal plants. Content of phenolic substances was determined by method of micellar electrokinetic capillary chromatography (MECC) and liquid chromatography (HPLC) in collection of 8 medicinal plants usually used in Czech Republic. For analysis was used freeze-dried and dried plant material. In freeze-dried material was found the highest content of total quercetin in Filipendula ulmaria L. (14200 mg/kg of dry weight) and in Betula pendula Roth. (11800 mg/kg of dry weight). The highest content of rutin contained Sambucus nigra L. (17700 mg/kg of dry weight). Similar values was measured in dried plant material. The content of total quercetin and rutin during drying was unchanged.
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